The overall objective of this proposal is to investigate 1) the role of DNA repair in chemotherapy resistance, and 2) the efficacy of bone-marrow-directed chemprotection in modulating anticancer chemotherapy. Specifically, the MGMT inhibitor O6- benzylguanine (BG) will be employed to selectively sensitize xenograft tumors to BCNU while simultaneously using retroviral transduction of MGMT mutants to protect bone marrow cells against BCNU-induced myelotoxicity. The point mutants MGMTP140A and MGMTP140K have been shown to be resistant to BG-induced sensitization to BCNU. The first specific aim is to develop an in vivo BG treatment regimen that inhibits wild-type MGMT in xenograft tumor cells for 24 hr. Xenograft human glioma SF767 tumors will be implanted s.c. into NOD/SCID mice, and various BG treatments will be compared. The second specific aim is to demonstrate the ability of transduced MGMT mutants to protect human cells from BG and BCNU combination therapy in vivo. Xenograft tumors of HeLa-mer (methylation repair deficient) cells stably expressing wild-type MGMT, MGMTP140A or MGMGP140K will be treated with the determined BG regimens in combination with BCNU. Resistance to BG-induced MGMT inhibition and BG + BCNU toxicity will be compared in vivo. The third specific aim is to investigate whether combination BG + BCNU chemotherapy can be made selectively cytotoxic to xenograft SF767 tumors in NOD/SCID mice transplanted with MGMTP140A- or MGMTP140K-transduced human bone marrow. MGMT inhibition, tumor regression, myelotoxicity, and other organ toxicities will be evaluated.